Compared to other gene transfer systems, retroviral vectors offer a wide range of advantages, including their ability to transduce a variety of cell types, to stably integrate transferred genetic material into the genome of the targeted host cell, and to express the transduced gene at significant levels. Therefore, vectors derived from the gamma-retroviruses, for example, the murine leukemia virus (MLV), have become a standard tool for gene transfer technology and have been frequently used in clinical gene therapy trials (Ross et al., Hum. Gen Ther. 7:1781-1790, 1996). Furthermore, the use of these vectors for the treatment of different monogenetic immunodeficiencies has also been previously described.
Pseudotyping of retroviral vectors, including HIV vectors or MLV vectors, refers to the incorporation of envelope proteins from heterologous viruses into the retroviral envelope membrane. Such pseudotyped retroviral vectors then exhibit a receptor phenotype similar to the virus from which the envelope protein was derived. Depending on the host range of said virus, the pseudotyped retroviral vectors will then have a broadened or a narrowed host range as compared to vector particles having the incorporated homologous retroviral envelope proteins.
A number of pseudotyped vectors have been described, including MLV vectors pseudotyped with the HIV Env protein, the Ebola virus glycoprotein, or the baculovirus glycoprotein. In all of these known vectors a single envelope protein was utilized to achieve the desired pseudotyping.
The measles virus (MeV), a prototype morbillivirus of the genus Paramyxoviridae, utilizes two envelope glycoproteins (the fusion protein (F) and the hemagglutinin protein (H)) to gain entry into the target cell. Protein F is a type I transmembrane protein, while protein H is a type II transmembrane domain, i.e. its amino-terminus is exposed directly to the cytoplasmic region. Both proteins thus comprise a transmembrane and a cytoplasmic region. One known function of the F protein is mediating the fusion of viral membranes with the cellular membranes of the host cell. Functions attributed to the H protein include recognizing the receptor on the target membrane and supporting F protein in its membrane fusion function. The direct and highly efficient membrane fusion at the cellular surface membrane is a particular property of measles virus and the morbilliviruses, thus distinguishing themselves from many other enveloped viruses that become endocytosed and will only fuse upon pH drop upon endocytosis. Both proteins are organized on the viral surface in a regular array of tightly packed spikes, H tetramers, and F trimers (Russell et al., Virology 199:160-168, 1994).
The Edmonton strain of MeV (MeVEdm) uses a single protein as its main receptor, namely, the protein known to be the regulator of complement activation factor, CD46 (Gerlier et al., Trends Microbiol. 3:338-345, 1995). CD46 is expressed on all nucleated human cells. Most clinical isolates of measles virus, however, can not effectively use CD46 as a receptor. Human SLAM (signaling lymphocyte-activation molecule; also known as CDw150) is a recently discovered membrane glycoprotein that is expressed on some T and B cells, and was also found to act as a cellular receptor for MeV, including the Edmonston strain (Tatsuo et al, Nature 406(6798):893-7, 2000).
The precise biological functions and interactions of the MeV H and F proteins remain largely unclear. However, in experiments where truncated F and H proteins were incorporated into recombinantly produced MeV vector particles (rMeV), Moll et al. (J. Virol. 76:7174-7186, 2002) concluded that rMeV containing the truncated F protein did not reveal any difference with wild type MeV (wt MeV) with respect to the induction of syncytium formation or in infectious-virus production. The rMeV containing the truncated H protein, however, exhibited a severely reduced induction of syncytium formation and a decrease in infectious-virus production. Thus, truncation of the F and H proteins differentially resulted in a decrease of titers or no effect at all.
In view of the importance of pseudotyped retroviral vectors for numerous gene transfer technology and clinical applications, there is still a great need for additional and improved pseudotyped retroviral vectors and methods for generating and using such vectors.